Two approaches are proposed to improve our understanding of eukaryotic gene expression and organization. The first aims to understand the organization of the Drosophila Gart locus, a gene encoding enzymatic activities for purine de novo synthesis. Our discovery of a cuticle protein gene within an intron of this gene raises the question of whether this arrangement has functional significance. We will carry out precise tests of various possibilities using germ-line transformation of in vitro-generated deletions. The first sequences that we will be testing are the highly conserved regions that are found throughout the cuticle protein gene region, entirely within an intron of the purine pathway gene. These extensive regions, found in a species comparison study, are thought to be involved in transcriptional regulation of the genes at the locus. Secondly, we hope to study other loci in the pathway for purine de novo synthesis, both because of our interest in this central metabolic pathway, and because of our hope that the complex gene organizational features found for Gart will thus be better understood. A novel approach will be used to isolate some of these genes. cDNA libraries will be constructed, and clones that represent the rare mRNA class at all stages will be used for very limited DNA sequence analysis. The resulting sequences will be conceptually-translated and compared to existing protein sequences which are known for nearly all of the purine pathway enzymes in various microbial organisms. As we will be accumulating a database which samples a large fraction of the "housekeeping" genes from Drosophila, we will also examine existing database sequences for protein sequence homologies. It is likely that some of these other genes that we identify by this approach will correspond to loci that were previously identified by mutation, thus furthering our understanding of the relationship between phenotype and gene function.